title: A methodology for radiolabeling of the endocannabinoid 2-arachidonoylglycerol (2-AG).
abstract: The metabolic intermediate and endocannabinoid signaling lipid 2-arachidonoylglycerol (2-AG) has not been readily labeled, primarily because of its instability toward rearrangement. We now detail a synthetic method that easily gives tritiated 2-AG from [5,6,8,9,11,12,14,15-(3)H(N)]arachidonic acid in two steps. We utilized a short chain 1,3-diacylglycerol and proceeded through the "structured lipid" [5'',6'',8'',9'',11'',12'',14'',15''-(3)H(N)]2-arachidonoyl-1,3-dibutyrylglycerol, a triacylglycerol that was conveniently deprotected in ethanol with acrylic beads containing Candida antarctica lipase B to give [5'',6'',8'',9'',11'',12'',14'',15''-(3)H(N)]2-arachidonoylglycerol ([(3)H]2-AG). The flash chromatographic separation necessary to isolate the labeled 2-acylglycerol [(3)H]2-AG resulted in only 4% of the rearrangement byproducts that have been a particular problem with previous methodologies. This reliable "kit" method to prepare the radiolabeled endocannabinoid as needed gave tritiated 2-arachidonoylglycerol [(3)H]2-AG with a specific activity of 200 Ci/mmol for enzyme assays, metabolic studies, and tissue imaging. It has been run on unlabeled materials on over 10 mg scales and should be generally applicable to other 2-acylglycerols.
authors.affiliation: Department of Pharmaceutical Sciences, Northeastern University, 360 Huntington Avenue, Boston, Massachusetts 02115, USA. duclos@neu.edu
authors.forename: Richard I
authors.initials: RI
authors.lastname: Duclos
authors.forename: Meghan
authors.initials: M
authors.lastname: Johnston
authors.forename: Subramanian K
authors.initials: SK
authors.lastname: Vadivel
authors.forename: Alexandros
authors.initials: A
authors.lastname: Makriyannis
authors.forename: Sherrye T
authors.initials: ST
authors.lastname: Glaser
authors.forename: S John
authors.initials: SJ
authors.lastname: Gatley
mesh_terms: D016718:Arachidonic Acid / Q000737:chemistry*; D001095:Arachidonic Acids / Q000737:chemistry*; D063385:Cannabinoid Receptor Modulators / Q000737:chemistry*; D004075:Diglycerides / Q000737:chemistry*; D063388:Endocannabinoids; D057075:Enzyme Assays / Q000379:methods; D005656:Fungal Proteins; D005989:Glycerides / Q000737:chemistry*; D007553:Isotope Labeling; D008049:Lipase / Q000737:chemistry*; D015394:Molecular Structure; D011869:Radioligand Assay; D015398:Signal Transduction
chemical_list: D001095:Arachidonic Acids; D063385:Cannabinoid Receptor Modulators; D004075:Diglycerides; D063388:Endocannabinoids; D005656:Fungal Proteins; D005989:Glycerides; D016718:Arachidonic Acid; C094503:glyceryl 2-arachidonate; D008049:Lipase; C431349:lipase B, Candida antarctica